The zinc-binding domain of mammalian prolyl-tRNA synthetase is indispensable for catalytic activity and organism viability

Kommireddy Vasu; Iyappan Ramachandiran; Fulvia Terenzi; Debjit Khan; Arnab China; Krishnendu Khan; Aayushi Chechi; Camelia Baleanu-Gogonea; Valentin Gogonea; Paul L Fox

doi:10.1016/j.isci.2021.102215

The zinc-binding domain of mammalian prolyl-tRNA synthetase is indispensable for catalytic activity and organism viability

iScience. 2021 Feb 20;24(3):102215. doi: 10.1016/j.isci.2021.102215. eCollection 2021 Mar 19.

Authors

Affiliations

¹ Department of Cardiovascular and Metabolic Sciences, Lerner Research Institute, Cleveland Clinic Foundation, Cleveland, OH, USA.
² Department of Chemistry, Cleveland State University, Cleveland, OH, USA.

Abstract

Aminoacyl-tRNA synthetases (AARS) participate in decoding the genome by catalyzing conjugation of amino acids to their cognate tRNAs. During evolution, biochemical and environmental conditions markedly influenced the sequence and structure of the 20 AARSs, revealing adaptations dictating canonical and orthogonal activities. Here, we investigate the function of the appended Zn²⁺-binding domain (ZBD) in the bifunctional AARS, glutamyl-prolyl-tRNA synthetase (GluProRS). We developed GluProRS mutant mice by CRISPR-Cas9 with a deletion of 29 C-terminal amino acids, including two of four Zn²⁺-coordinating cysteines. Homozygous ZBD mutant mice die before embryonic day 12.5, but heterozygous mice are healthy. ZBD disruption profoundly reduces GluProRS canonical function by dual mechanisms: it induces rapid proteasomal degradation of the protein and inhibits ProRS aminoacylation activity, likely by sub-optimal positioning of ATP in the spatially adjacent catalytic domain. Collectively, our studies reveal the ZBD as a critical determinant of ProRS activity and GluProRS stability in vitro and in vivo.

Keywords: Biochemistry; Biological Sciences; Molecular Biology.